Project Summary The goal of this project is to gain deep mechanistic understanding and predictive capability of the molecular mechanisms that govern transport through the mucus barrier. The mucus lining forms a selective barrier that facilitates the uptake of nutrients, sperm, and oxygen, while preventing free passage of harmful viruses, bacteria, and toxins. The detailed molecular properties that distinguish particles that pass through the mucus barrier and particles that are rejected by the mucus barrier are largely unknown, and hence, predictive models for mucosal transport are currently missing, despite their tremendous implications for drug delivery and preventing prevalent infectious diseases (for example Papilloma virus, HIV). While the relevance of particle size, net charge, and hydrophobicity for mucus transport has been studied in isolation, the effect of combining these properties and the role of spatial arrangement have not been studied in a way that allows to predict mucus-interactions or to design drug delivery vehicles with tailored mucus transport properties. We propose to characterize molecular transport through the mucus barrier and relate the results to the spatial surface arrangement of charge, hydrophobicity and specific peptide sequences. This knowledge will enable us to determine biophysical fingerprints that are diagnostic for fast and slow passage, and has the potential to transform the design of drug delivery vehicles as it will allow to combine surface functionalization (for tissue targeting) while independently tuning the transport properties of a vehicle. In the first aim we will test the influence of charge distribution and hydrophobicity for transport through mucus using short peptides with systematically varied residues, and a microfluidic system to measure uptake, spatial distribution, and transport through the mucus. In the second aim, we will use phage-display-based approaches to determine whether these same rules, when applied to peptides on the surface of a particle (specifically, phage) can facilitate passage of the particle through mucus. This system will also give insight into other parameters that affect particle-mucus interactions, such as peptide length, specific residue sequence, surface display density, and particle geometry. In Aim 3 we will integrate the knowledge from Aims 1 and 2 and determine the relevance of surface charge, hydrophobicity, and specific peptide sequences influences mucus transport in vivo, using the mouse vagina as a model. The multidisciplinary research team presents the expertise necessary for combining fundamental science questions with cutting edge engineering applications: a biologist with experimental and theoretical expertise in biological hydrogel systems, a mechanical engineer with expertise in transport phenomena in biological tissues, a synthetic biologist with expertise in engineering phage display systems, and a chemical engineer with expertise in controlled particle surface functionalization and characterization in vivo.